Doctor Dr. Prithviraj Nandigrami "Raj"
Biochemistry/TCB
UIUC
Urbana, IL

Monday, November 27, 2017
3:00 pm (CT)
3269 Beckman Institute

Abstract

Conformational dynamics is often essential for a protein’s function. For example, proteins are able to communicate the effect of binding at one site to a distal region of the molecule through changes in its conformational dynamics. This so called allosteric coupling fine tunes the sensitivity of ligand binding to changes in concentration. A conformational change between a “closed” (apo) and an “open” (holo) conformation upon ligation often produces this coupling between binding sites. Enhanced sensitivity between the unbound and bound ensembles leads to a sharper binding curve. In this work, I focus on molecular dynamics simulations to understand microscopic origins of ligand binding cooperativity using a ubiquitous calcium-binding protein Calmodulin (CaM). Domain opening transitions of isolated domains of CaM in the absence of calcium shows that the simulated transition mechanism of nCaM follows a two-state behavior, while domain opening in cCaM involves global unfolding and refolding of the tertiary structure. The unfolded intermediate also appears in the landscape of nCaM, but at a higher temperature than it appears in cCaM’s energy landscape. I also investigate the structural origins of binding affinity and allosteric cooperativity of binding two calcium ions to each domain of CaM. I analyze the simulated binding curves within the framework of the classic Monod-Wyman-Changeux (MWC) model of allostery to extract the binding free energies to the closed and open ensembles. The analysis of the simulations offers a rationale for why the two domains differ in cooperativity: the higher cooperativity of cCaM is due to larger difference in affinity of its binding loops. Finally, I extend the work to investigate structural origins of binding cooperativity of four calcium ions to intact CaM. I focus on investigating the influence of this heterogeneity on the kinetic flux of binding pathways as a function of concentration. The formalism developed for Network Models of protein folding kinetics is used to evaluate the directed flux of all possible pathways between unligated and fully loaded CaM.